Many commonly used anti-cancer drugs indiscriminately target DNA in dividing cells and ultimately cause DNA damage. This, in turn, triggers activation of cell cycle checkpoints which arrest progression of the cell cycle (at the G1, S, or G2/M phases) with the purpose of allowing time for the DNA to be repaired before the cell undergoes DNA replication or division. From a therapeutic standpoint, inhibition of checkpoint kinases that mediate cell cycle arrest could force tumor cells to continue cell division before chemically-induced DNA damage is repaired, eventually causing apoptosis or mitotic catastrophe (Medema, R. H. and Macurek, L., Oncogene, 2012, 31(21):2601-2613). Cell line studies support this hypothesis and show chemosensitization and radiosensitization by pharmacologic or genetic disruption of checkpoint kinase activity including CHK1, WEE1, ATR, and ATM. Inhibitors against these kinases are at various stages of preclinical and clinical development for their ability to sensitize tumor cells to therapeutic DNA damage.
The checkpoint kinase WEE1 catalyzes an inhibitory phosphorylation of both CDK1 (CDC2) and CDK2 on tyrosine 15 (Parker, L. L. and Piwnica-Worms, H., Science, 1992, 257(5078):1955-1957; Watanabe, N., et al., Embo J., 1995, 14(9):1878-1891). WEE1-dependent inhibition of CDK1 and CDK2 arrests the cell cycle in response to extrinsically induced DNA damage (Hamer, P. C. D., et al., Clin. Cancer Res., 2011, 17(13):4200-4207). WEE1 activity is also essential for the unperturbed cell cycle (Mcgowan, C. H. and Russell, P., Embo J., 1993, 12(1):75-85; Tominaga, Y., et al., Intl. J. Biol. Sci., 2006, 2(4):161-170). Cell synchronization studies in normal human fibroblasts revealed that similar amounts of WEE1 protein were detected in both S and G2/M phases, but that its greatest activity was in S phase of the cell cycle (Watanabe, N., 1995). Further, upon conditional WEE1 knockout in mouse embryonic fibroblasts (MEFs), cells show evidence of genomic instability, malfunctioning checkpoints, and premature mitosis (Tominaga, et al., 2006). This phenotype was explained in part by recent findings that demonstrate a critical role for WEE1 in DNA synthesis. Knockdown of WEE1, in the absence of DNA damaging agents, led to rapid and robust detection of DNA double strand breaks specifically in S-phase cells undergoing DNA replication (Beck, H., et al., J. Cell Biol., 2010, 188(5):629-638; Dominguez-Kelly, R., et al., J. Cell Biol., 2011, 194(4):567-579). Data support a model of WEE1-dependent genomic stability in which WEE1 knockdown or inhibition leads to aberrantly high activity of CDK 1 and 2, resulting in inappropriately timed firing of excessive DNA replication origins that quickly depletes nucleotide pools and leads to stalled replication forks which, in the absence of WEE1 activity, are substrates for DNA exonucleases and resolve into DNA doubles strand breaks (Beck, H., et al., 2012).
Deregulated WEE1 expression or activity is believed to be a hallmark of pathology in several types of cancer. WEE1 is often overexpressed in glioblastomas and its activity protects this tumor type from mitotic catastrophe such that high WEE1 levels are associated with poor prognosis (Mir, S. E., et al., Cancer Cell, 2010, 18(3):244-257). High expression of WEE1 was found in malignant melanoma and correlated with poor disease-free survival in this population (Magnussen, G. I., et al., Plos One, 2012, 7(6)). Aberrant WEE1 expression has been implicated in additional tumor types such as hepatocellular carcinoma (Masaki, T., et al., Hepatology, 2003, 37(3):534-543), breast cancer (Iorns, E., et al., Plos One, 2009, 4(4)), colon carcinoma (Backert, S., et al., Intl., J. Cancer, 1999, 82(6):868-874)), lung carcinoma (Yoshida, T., et al., Annals of Oncology, 2004, 15(2):252-256) and head and neck squamous cell carcinoma (Wu, Z. X., et al., Mol. & Cell. Proteomics, 2011, 10(12)). Advanced tumors with an increased level of genomic instability may require functional checkpoints to allow for repair of such lethal DNA damage. As such, WEE1 represents an attractive target in advanced tumors where its inhibition is believed to result in irreparable DNA damage (Sorensen, C. S. and Syljuasen, R. G., Nuc. Acids Res., 2012, 40(2):477-486).
There is a need for biomarkers that can be used to predict which patients are amenable to treatment with specific therapies, particularly for patients who are non-responsive or who are likely to become refractive to first line therapies. It is, therefore, an object of this invention to provide a predictive biomarker to select patients likely to respond to treatment with a WEE1 inhibitor.